The studies of this application ask how gene expression controls neuronal connectivity and, thereby, brain function. This question is of general importance if one wants to understand, and (therapeutically) manipulate brain circuitry. We use the model system Drosophila, where virtually every gene can be targeted for knock-out or activation in a cell type selective manner Drosophila also offers the advantage that its brain i composed of a relatively small number of stereotyped neuronal lineages, groups of neurons descended from individual embryonic stem cells, called neuroblasts. During the course of its proliferation, each neuroblast expresses characteristic sets of regulatory genes. These genes control the differentiation of the neurons born from that particular neuroblast during a particular time interval. Through this mechanism, a lineage, or smaller subdivision of a lineage called sublineage, develops into a specific class of neurons which share common wiring properties, including the projection of their axons, branching pattern, and placement of synapses. Several discrete neuronal classes/lineages are put together into a neuronal circuit. We have identified a circuit, called the anterior visual pathway (AVP), which conducts input form the eye to a brain center, the central complex, known to process and store visual information in order to control fly locomotion (walking, flight). The central part of this circuit is formed by three lineages, whose neurons form several classes of highly ordered parallel and sequential elements. In our first aim we will investigate the function of the neuronal classes of the AVP, by recording their activity in response to defined visual stimuli. We will also demonstrate experimentally that these classes of neurons are directly connected by synapses. The second aim addresses the question how the developmental history of a neuron (time of birth, placement within the spatial framework of the developing brain) relates to its later connectivity within the AVP circuit. Furthermore, by genetically ablating specific classes of AVP neurons and monitoring the response of their normal synaptic partners, we will obtain important clues towards the role of specific cell interactions ordering connectivity. Thirdly, using high throughput RNAseq, we will analyze the complete assortment of genes (transcriptome) expressed differentially in two particular AVP sublineages, R3 and R2. These two classes are distinguished from each other by very few structural criteria, and we will screen for and then analyze genes responsible for their differences in wiring. We expect to identify genes which play a general role in controlling pathway choices and connectivity in the nervous system.